Researchers can now get bacteria to run a major biochemical pathway in reverse …

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The majority of plant matter we have available to produce biofuels comes in the form of cellulose, a long polymer of sugars. It's easiest to convert this material to ethanol, but that creates its own problems: ethanol is less energy dense than petroleum-based fuels, and most vehicles on the road can't burn more than a 15 percent mix of ethanol and standard gasoline.

These disadvantages have led a number of labs to look into ways of using a cellulose feedstock to produce something more like standard fuels. In yesterday's Nature, researchers proposed a clever way of doing this: take the biochemical pathway that normally burns fat and run it in reverse.

Not just one-way

Cells have a pathway for the production of fatty acids, long hydrocarbon chains that are normally linked together to form fats. The end products at least look a bit more like the fuels that currently run our cars than ethanol does, but using this pathway to produce biofeuls has drawbacks. It requires a substantial input of energy in the form of ATP and tends to produce hydrocarbon chains that are too long (10-20 carbons long) to make a really convenient fuel. This pathway is also tightly regulated, since most microbes would rather divert their energy to reproduction than to making fat.

As a result, a team of researchers from Rice University decided to forgo this pathway entirely. They reasoned that cells have a second, entirely separate set of enzymes normally used to break fats down that might be repurposed to make biofuel.

Enzymes are catalysts. They generally act by making a chemical reaction more likely to occur—they don't usually dictate in which direction the reaction goes. So, if you supply an enzyme with a large quantity of what are normally the end products of a given reaction, it will readily catalyze the reverse reaction. If you run the pathway that normally digests fats in reverse, it will produce longer hydrocarbons.

Sounds simple, right? But actually getting bacteria (the authors worked with E. coli) to do this isn't necessarily easy. To begin with, the bacteria won't produce any of these necessary enzymes unless they think they have fat to digest. Years of genetic studies have identified the genes responsible for shutting off the fat burning pathway, so the authors knocked those genes out.

Problem solved? Not quite. Even when fat is available, E. coli would rather burn simple sugars instead if they're present. The gene that mediates this preference has also been identified, and the authors spliced a mutant form of it into the bacteria's DNA. WIth these mutations in place, the bacteria would at last have the right enzymes around, no matter what the conditions.

The authors fed their modified E. coli glucose, which can be produced by the breakdown of cellulose (meaning the process is biofuel compatible). Glucose is a six-carbon molecule that's broken down into short, two-carbon chunks in a process that produces ATP to fuel the cell. These two carbon molecules end up attached to a co-factor in a molecule called acetyl-Coenzyme A. If oxygen is present, acetyl-CoA gets handed over to a process that produces a number of ATP molecules as acetyl-CoA is converted into water and carbon dioxide (the CoA is recycled). If oxygen is not present, organisms like yeast convert acetyl-CoA into ethanol instead, freeing up the CoA for reuse.

As it turns out, acetyl-CoA is also where the digestion of fats feeds into the normal metabolism. So, by giving the bacteria lots of glucose, the authors created conditions where the end product of fat digestion, acetyl-CoA, was present in abundance, but there wasn't an excess of the starting material, namely fat. This was enough to tip the pathway backwards, building up longer chains of hydrocarbons. To give the system an extra boost, the authors knocked out the gene that sends acetyl-CoA down the pathway towards ethanol.

On its own, this process wouldn't do anything useful, since it would create a mix of longer hydrocarbons all linked up to coenzyme A. But organisms have ways of diverting specific products for use in the production of specific molecules they need, such as amino acids or the bases of DNA. So the authors did a bit more engineering and added some copies of the gene that divert a four-carbon intermediate into butanol. Expression of a different gene shifted the production toward longer hydrocarbons, resulting in a mix of molecules that contain a chain of 12 to 18 carbon atoms. Almost all of the reactions researchers tested resulted in the most efficient production of end products that anyone has reported.

So much potential

There's so much to like in this paper. To begin with, the authors are successfully leveraging decades of bacterial genetics and basic biochemistry to do this work. They really are building something using information that was pieced together by hundreds of researchers, most of whom probably didn't ever think their work would have implications for the oil economy.

It's also simply a tour de force of genetic engineering. Every time a reaction went too slowly, the researchers would pop a few extra copies of the relevant genes in to speed it up. Any sign of unwanted byproducts and they knocked out the genes that produced them.

There's a tremendous amount of potential here. The authors have shown that it's possible to divert this pathway into a variety of products, but they've only done so by altering a limited number of genes, generally the ones that already exist in E. coli. There's a whole world of other bacteria out there, so it may be possible to identify genes that can use the same process to create a huge array of other useful products.

But, perhaps more significantly, the pathway is generally helpful to the cell, in that it acts in much the same way that ethanol production does when the bacteria are deprived of oxygen: it gets some ATP made from glucose and allows the cell to recycle key components of its metabolism. In this way, it avoids the biggest problem with many biofuels, namely that the energetic cost of producing them provides a selective pressure for the cells to evolve ways of disabling the pathway. In fact, since the cells can rely on this pathway for ATP production, this approach may even induce them to evolve ways of making it more efficient.

Awesome, it's like an alternative War of the Worlds - Big Oil taken down by E. coli.

Actually, Big Oil has the capital, expertise, and infrastructure to take a technology like this and exploit it better than almost anyone else can. If this can make them money, then they will invest in it and improve it, count on it.

So, can they do the same thing to convert celulose to glucose? As I understand it, that's still an energetically expensive part of the process for forming ethanol.

The process of converting cellulose to glucose is now generally done with enzymes from certain microbes. Imagine if they could genetically engineer two symbiotic species, one eating cellulose and secreting glucose, the other eating the glucose and secreting gasoline-like molecules (like the E.Coli in this paper). It may even be possible to combine both function into the same cell, allowing us to produce viable biofuel directly from cellulose.

Awesome, it's like an alternative War of the Worlds - Big Oil taken down by E. coli.

Actually, Big Oil has the capital, expertise, and infrastructure to take a technology like this and exploit it better than almost anyone else can. If this can make them money, then they will invest in it and improve it, count on it.

Great article Ars. This really is an exciting discovery and has the potential to be as important to transport as the Haber Bosch process is to agriculture. It could even help reduce the CO2 content of the atmosphere - suck the CO2 out with this process, and pump the hydrocarbons back into old oil wells. I guess that might be getting a bit ahead of ourselves.

The best part is how they made evolution work for them to make the process more efficient, even though evolution doesn't exist. Go figure.

One slightly worrisome thing here is that by engineering a pathway that is advantageous to the organism, it would seem that would raise the likelyhood that the organism could survive on its own in the wild, i.e. danger of uncontrolled propagation from any accidental release. How unique is this? I have always thought that engineered organisms were almost universally less fit than their wild progenitors, both by design and by interfering in a selection landscape the details of which we have only glimmers of understanding.

One slightly worrisome thing here is that by engineering a pathway that is advantageous to the organism, it would seem that would raise the likelyhood that the organism could survive on its own in the wild, i.e. danger of uncontrolled propagation from any accidental release. How unique is this? I have always thought that engineered organisms were almost universally less fit than their wild progenitors, both by design and by interfering in a selection landscape the details of which we have only glimmers of understanding.

Flesh-eating bacteria -> fat producing bacteria?

And thus was born the Blob.

Advantageous to the organism in the strange environment of the cellulose dissolving tank (kept at the ideal temperature, pressure, humidity, and pH for the production of what we want).

Any that escape face the real world, which is considerably less ideal, incredibly varied, and densely populated with their wild cousins who have not had monkeys smashing up their finely balanced billion year perfected chemical engines.

Remember, the researchers did a lot of tinkering, but it was all in the context of "break this thing we don't want it to do anymore, and put in more copies of the thing we want it to do"... random mutation and copying errors do that ALLL the time, it just doesn't stick unless it is useful (and there aren't many cellulose breakdown vats outside of the deep sea pressure zones that created our fossil fuels).

Even better would be a symbiotically pairing a photosynthesizing organism that produces glucose with this kind of bacteria. You could have whole farms of this stuff just sitting out in the sun oozing out something like fuel oil. Hell, you could pair any organism that produces glucose from any kind of energy source with something like this, and have a really versatile tool for energy storage. Hmm, you would want to make sure it is restricted some how to a man-made environment, would't want it getting out all over into the environment.

Awesome, it's like an alternative War of the Worlds - Big Oil taken down by E. coli.

Actually, Big Oil has the capital, expertise, and infrastructure to take a technology like this and exploit it better than almost anyone else can. If this can make them money, then they will invest in it and improve it, count on it.

One can only hope they will.

Not really. While I agree with the assertion that Big Oil *could* do the most with this technology, I can all but guarantee you that their doing so will only make gas prices higher. They'll justify the price boost by claiming research costs, production issues, etc, and get everyone to buy into it by playing the environmental card, but the net result would be another irreversible hike in gas prices.

The sad thing is that Rice is in Houston and Big Oil is heavily involved with the university. Conspiracy theorists don't have to make a crazy stretch to claim that this work will be quietly suppressed. I sure hope Rice will license this technology to entities that want to make it commercially successful.

The sad thing is that Rice is in Houston and Big Oil is heavily involved with the university. Conspiracy theorists don't have to make a crazy stretch to claim that this work will be quietly suppressed.

Actually, yes, they do.

Oil is becoming increasingly harder to pump out of the ground, and increasingly more expensive as a result. Big oil companies have a lot of money invested in their oil wells, some of which is recoverable when the oil runs "dry" (meaning it costs more to extract than it can be sold for), so they're not keen to lose them ... but they also have a lot of money invested in refineries and other infrastructure for shipping oil around the world.

If they manage to scale this up to the point where they can produce a barrel of oil for (say) $US50, and do so reliably, they'll stop spending money on exploration and developing new sites, especially if those new sites cost more than $US50 a barrel to extract the oil, and they'll use this process instead.

It's a very simple equation. Companies exist to make money. They'll resist a disruptive technology as long as they can, if they have the power to do so, but sooner or later, if this technique is as good as the writeup implies, it'll win. The basic information is out there now; the genie can't be put back into the bottle.

It's a very simple equation. Companies exist to make money. They'll resist a disruptive technology as long as they can, if they have the power to do so, but sooner or later, if this technique is as good as the writeup implies, it'll win. The basic information is out there now; the genie can't be put back into the bottle.

I don't think anyone doubts that big companies will use this tech eventually... I just think there is doubt that they will move their asses and get to it in our lifetimes. Sure, f(company)=$... but there is a massive damping on that function, more powerful than greed, called laziness. The scientists working on this project are probably just going to get paid a decent hunk of money to go study something else, and this research will be sat on until the last extant whale gets forked from the ocean for its oil.

Awesome, it's like an alternative War of the Worlds - Big Oil taken down by E. coli.

Actually, Big Oil has the capital, expertise, and infrastructure to take a technology like this and exploit it better than almost anyone else can. If this can make them money, then they will invest in it and improve it, count on it.

This is the truth. Everyone likes to cry and moan about "Big Evil oil" forgetting they have revolutionized the way we live. They dont want to suppress or block new energy technologies when they could put those into production. More importantly, as you say the only ones with the infrastructure and capital to do that are probably those big evil oil companies that throw baby kittens into furnaces to make that black sticky oil.....

I don't think anyone doubts that big companies will use this tech eventually... I just think there is doubt that they will move their asses and get to it in our lifetimes. Sure, f(company)=$... but there is a massive damping on that function, more powerful than greed, called laziness. The scientists working on this project are probably just going to get paid a decent hunk of money to go study something else, and this research will be sat on until the last extant whale gets forked from the ocean for its oil.

Peak oil. http://en.m.wikipedia.org/wiki/Peak_oil. It will get more expensive over time. And become more environmentally destructive, such as the oil sands. The more the devastation, the more they spend fighting off stronger regulation. There's no way they are going to ignore this, they will at least quietly invest in the technology. Once cheap enough, they'll abandon drilling and capture from the earth, assuming you can safely and efficiently generate fats in this manner.

Regarding whales, we already did this. It's called the 19th century, infamously immortalized in Moby Dick. They had quite the process back then. Problem is, those whales get scarce quite fast, hence the switch to petroleum well before the rise of the automobile. After all, how many cars were on the road while Pennsylvania was the lead producer in the 1870's? Gasoline was actually an undesirable product of oil refinement for lubrication and fuels, because it was too volatile. Which turned out to be just what the internal combustion engine needed.

The sad thing is that Rice is in Houston and Big Oil is heavily involved with the university. Conspiracy theorists don't have to make a crazy stretch to claim that this work will be quietly suppressed.

Actually, yes, they do.

Oil is becoming increasingly harder to pump out of the ground, and increasingly more expensive as a result. Big oil companies have a lot of money invested in their oil wells, some of which is recoverable when the oil runs "dry" (meaning it costs more to extract than it can be sold for), so they're not keen to lose them ... but they also have a lot of money invested in refineries and other infrastructure for shipping oil around the world.

If they manage to scale this up to the point where they can produce a barrel of oil for (say) $US50, and do so reliably, they'll stop spending money on exploration and developing new sites, especially if those new sites cost more than $US50 a barrel to extract the oil, and they'll use this process instead.

It's a very simple equation. Companies exist to make money. They'll resist a disruptive technology as long as they can, if they have the power to do so, but sooner or later, if this technique is as good as the writeup implies, it'll win. The basic information is out there now; the genie can't be put back into the bottle.

All they have to do is buy them out and patent it and there you go. Genie back in bottle. This isn't their first 'buying out the alternative energy tech' rodeo.

Awesome, it's like an alternative War of the Worlds - Big Oil taken down by E. coli.

Actually, Big Oil has the capital, expertise, and infrastructure to take a technology like this and exploit it better than almost anyone else can. If this can make them money, then they will invest in it and improve it, count on it.

Apple laptops powered by your friendly nearby e.coli, just feed it Jack in the Box. After all.. Apple has larger market cap than even exxon mobile

This paper sounds like it heralds two separate groundbreaking pieces of science: genetically engineered bacteria and the really clever insight that on the cellular level, inputs and outputs can be reversed! It sounds like very promising subfields of biology and engineering have just been opened.

With that weight in mind, I thought I would comment on the particular application they chose. Crude oil ends up in far more places than internal combustion engines and it sounds like all the other users probably couldn't use pure ethanol so this is also promising for them. I also feel like I should mention that this biofuel sounds like it will have all the drawbacks of crude oil except that it won't run out but making it is going to make food more expensive in a world that was already having trouble paying for it.

I'm personally an optimist so I hate to leave my comment on such a negative nitpick so I wanted to thank Ars in general and John Trimmer personally for this great article! The author's excitement really made this article great and I wouldn't understand a few things that I can see now without the excellent writing.

Now they just have to evolve the bacteria to make them more efficient. They should also evolve something that is unnaturally efficient at converting sunlight+water+CO2 into glucose and make them co-exist.